Yield and Quality of RNA from Grape Berries at Different Developmental Stages

Abstract

Studies of gene expression in grape berries require extraction of high quality RNA for construction of cDNA libraries and northern blot analyses. We have developed a procedure that yields high quality RNA from grape berries and have used it to extract RNA from fruit at five stages of development, from just after set until just after veraison. The extraction is based on a Tris-borate extraction buffer that has proven useful in other fruit tissue containing large amounts of polysaccharide. The method does not require cesium chloride centrifugation but includes a series of ethanol precipitations to remove polysaccharides. The quality of the RNA was demonstrated by constructing a cDNA library from a berry sample just prior to veraison. The library had 6 x 10⁵ primary recombinants with an average insert size of 1.3 kb. We were able to obtain several full length clones for malic enzyme from the library, one of which was sequenced and is available on Genbank (#L-34836).